| Abstract: | Enzyme stability and enzyme kinetic studies with
plasmid.mediated and chromosome-mediated beta-
lactamases against cefotaxime, cefoperazone and
other cephalosporins were performed using spectro-
photometry. Beta-lactamases were extracted from
granvnegative organisms and identified by isoelectric
focusing with the special reagent nitrocefin. The
relative hydrolysis rate (%), Michaelis-Menten constant
(Km), maximum velocity (Vmax) and inhibition
constant (Ki) were obtained and used as indicators
for comparative study of enzyme stability in six
cephalosporins. The affinity of compounds bindinp;
to target sites of beta-lactamases was also determined.
The results show that cefotaxime is the most
enzyme stable drug among the cephalosporins studied.
The Vmax of TEM-1, SHV-I and P99 enzyme hydro
lyzing cefotaxime was less than 0.10% as compared
with cephaloridine. Cefotaxime demonstrates high
affinity to the chromosmm>mediated enzyme P99,
and its Ki value is 0.036 µm. Cefoperazonc exhibits
less enzyme inhibitory activity and is unstable with
most of the beta-lactamases used. |
